| Monascus purpureus MQ-01 producing high monascus pigments in fujian was used as the starting strain,Genomiec DNA was extracted from M.purpureus MQ-01 by using an improved CTAB method.The primers were designed accroding to the gene coding for a key ketidesynthase of citrinin in Genebank,the genomic DNA as a template was used to amplify the gene coding for a key ketidesynthase of citrinin by PCR.The gene was inserted into pBluescript sk (+) DNA and transformed into E. coli DH5a.Sequencing the DNA sequence of transformants,the results showed that the gene was just the gene coding for a key ketidesynthase of citrinin.The constructed vector digested with restriction enzyme was linked into the trpC gene promoter and Hygromycin B resistance gene hph sequence of pCSN43 plasmid by polymerase chain reaction.Preparation of M.purpureus MQ-01 protoplasts at first,then M.purpureus MQ-01 was transformed by restriction enzyme-mediated transformation,finally the transformant M.purpureus MN-02 was screened by hygromycin B resistant.Compared levels of monascus pigments and citrinin contents of original strain MQ-01 with transformant MN-02 during solid-state fermentation and submerged fermentation,the results showed that:comparing with the original strain MQ-01,the levels of monascus pigments of transformant MN-02 were not reduced,and increased slightly during solid-state fermentation and submerged fermentation.Monascus pigments products were improved to 30.5 U/g and 4.05% during solid-state fermentation by transformant MN-02,and they were improved to 17.85 U/g and 10.53% during submerged fermentation. Citrinin contents of original strain MQ-01 were 32.875μg/g during during solid-state fermentation and submerged fermentation.However,citrinin couldn't be detected during solid-state fermentation and submerged fermentation by transformant MN-02 under the experimental conditions.Therefore,the experiment obtained the safe strain with lower citrinin contents even no citrinin. |
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